Organic compounds

ABSTRACT

The invention relates to a new crystalline form II of N-benzoyl-staurosporine; compositions containing the same; processes for the preparation thereof; and the use of crystalline form II of of N-benzoyl-staurosporine in diagnostic methods or therapeutic treatment of warm-blooded animals, especially humans. The invention relates to the amorphous forms of N-benzoyl-staurosporine; compositions containing the same; processes for the preparation thereof; and the use of amorphous N-benzoyl-staurosporine in diagnostic methods or therapeutic treatment of warm-blooded animals, especially humans.

This application is a Divisional of U.S. application Ser. No. 11/718,029filed Apr. 26, 2007 which is the National Stage of Application No.PCT/EP2005/0011789, filed on Nov. 3, 2005, which claims benefit under 35U.S.C. §119(e) of U.S. Provisional Application No. 60/625,343, filedNov. 5, 2004 and U.S. Provisional Application No. 60/642,131 filed Jan.7, 2005. The contents of which are incorporated herein by reference intheir entirety.

The invention relates to a new crystalline form II ofN-benzoyl-staurosporine, the process for preparation of the crystallineform II of N-benzoyl-staurosporine, compositions containing crystallineform II of N-benzoyl-staurosporine, and the use of crystalline form IIof N-benzoyl-staurosporine in diagnostic methods or therapeutictreatment of warm-blooded animals, especially humans.

The invention relates to amorphous N-benzoyl-staurosporine, the processfor the preparation of amorphous N-benzoyl-staurosporine, compositionscontaining amorphous of N-benzoyl-staurosporine, and the use ofamorphous N-benzoyl-staurosporine in diagnostic methods or therapeutictreatment of warm-blooded animals, especially humans.

BACKGROUND OF THE INVENTION

The drug N-benzoyl-staurosporine is used as an anti-tumour agent. Ingeneral, the preparation of N-benzoyl-staurosporine is known in the art.However, it is also known that different polymorphic forms of the samedrug may have substantial differences in certain pharmaceuticallyimportant properties. Therefore, there is a continuing need for newsolid forms of N-benzoyl-staurosporine and new methods of preparation.

SUMMARY OF THE INVENTION

In accordance with one aspect, the invention provides a crystalline formII of N-benzoyl-staurosporine. Preferably, the crystalline form II ofN-benzoyl-staurosporine has an X-ray diffraction pattern with a peak atan angle of refraction 2-theta (θ) of 8.7±0.2 as depicted in FIG. 1.

In accordance with yet another aspect, the invention provides acomposition that contains N-benzoyl-staurosporine in a solid form,wherein at least 80% by weight of the solid N-benzoyl-staurosporine isits crystalline form II having an X-ray diffraction pattern with a peakat an angle of refraction 2θ of 8.7±0.2 as depicted in FIG. 1. Variousembodiments and variants are provided.

In accordance with yet another aspect, the invention provides apharmaceutical composition that includes crystalline form II ofN-benzoyl-staurosporine and a pharmaceutically acceptable carrier ordiluent. Preferably, the pharmaceutical composition is for oraladministration.

In accordance with yet another aspect, the invention also relates toamorphous N-benzoyl-staurosporine, the process for the preparation ofamorphous N-benzoyl-staurosporine and compositions containing amorphousN-benzoyl-staurosporine.

In accordance with another aspect, the invention provides apharmaceutical composition that includes a prophylactically ortherapeutically effective amount of amorphous N-benzoyl-staurosporineand one or more pharmaceutically acceptable excipients. Thepharmaceutical compositions of this aspect of the invention may beformulated, e.g., for oral administration.

In accordance with yet another aspect, the invention provides processesfor purifying staurosporine.

In accordance with yet another aspect, the invention provides a processfor the preparation of N-benzoyl-staurosporine by reacting staurosporinewith benzoic anhydride. This process optionally further comprisesseeding the reaction solution to produce either amorphous or crystallineform II of N-benzoyl-staurosporine.

In accordance with yet another aspect, the invention provides a processfor the preparation of amorphous N-benzoyl-staurosporine involving spraydrying a solution containing N-benzoyl-staurosporine.

Pharmaceutical compositions that include a prophylactically ortherapeutically effective amount of amorphous N-benzoyl-staurosporineproduced by the process described, and one or more pharmaceuticallyacceptable excipients are also provided.

In accordance with yet another aspect, the invention provides a processfor making the crystalline form II of N-benzoyl-staurosporine, theprocess including:

-   -   (a) providing a solution of N-benzoyl-staurosporine in either a        protic or an aprotic solvent;    -   (b) seeding with the crystalline form II of        N-benzoyl-staurosporine and contacting the reaction mixture with        an alcohol solvent to form a precipitate; and    -   (c) isolating the precipitate, which is the crystalline form II        of N-benzoyl-staurosporine. Various embodiments and variants are        provided.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the X-ray powder diffraction diagram of crystalline form IIof N-benzoyl-staurosporine.

FIG. 2 shows a characteristic infrared spectrum of crystalline form IIof N-benzoyl-staurosporine.

FIG. 3 shows the X-ray powder diffraction diagram of a purely amorphousform of N-benzoyl-staurosporine.

FIG. 4 shows the X-ray powder diffraction diagram of an essentiallyamorphous form of N-benzoyl-staurosporine.

DETAILED DESCRIPTION OF THE INVENTION

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Although any methods andmaterials similar or equivalent to those described herein can be used inthe practice or testing of the present invention, the preferred methodsand materials are described.

For the purposes of the present invention, the following terms aredefined below.

The crystalline compound, designated herein as “crystalline form II” andreferred to hereinafter as crystalline form II ofN-benzoyl-staurosporine. It is characterized via X-ray powderdiffraction, and/or infrared spectroscopy. It is further describedbelow.

“Pharmaceutically acceptable” means that which is useful in preparing apharmaceutical composition that is generally non-toxic and is notbiologically undesirable and includes that which is acceptable forveterinary use and/or human pharmaceutical use.

“Anti-solvent” is a solvent which when added to an existing solution ofa substance reduced the solubility of the substance.

The term “composition” includes, but is not limited to, a powder, asolution, a suspension, a gel, an ointment, an emulsion and/or mixturesthereof. The term composition is intended to encompass a productcontaining the specified ingredients in the specified amounts, as wellas any product, which results, directly or indirectly, from combinationof the specified ingredients in the specified amounts. A “composition”may contain a single compound or a mixture of compounds. A “compound” isa chemical substance that includes molecules of the same chemicalstructure.

The term “pharmaceutical composition” is intended to encompass a productcomprising the active ingredient(s), pharmaceutically acceptableexcipients that make up the carrier, as well as any product whichresults, directly or indirectly, from combination, complexation oraggregation of any two or more of the ingredients, or from dissociationof one or more of the ingredients, or from other types of reactions orinteractions of one or more of the ingredients. Accordingly, thepharmaceutical compositions of the present invention encompass anycomposition made by admixing the active ingredient, additional activeingredient(s) and pharmaceutically acceptable excipients.

The term “excipient” means a component of a pharmaceutical product thatis not the active ingredient, such as filler, diluent and carrier. Theexcipients that are useful in preparing a pharmaceutical composition arepreferably generally safe, non-toxic and neither biologically norotherwise undesirable, and are acceptable for veterinary use, as well ashuman pharmaceutical use. “A pharmaceutically acceptable excipient”, asused in the specification and claims, includes both one and more thanone such excipient.

“Therapeutically effective amount” means the amount of a compound that,when administered for treating or preventing a disease, is sufficient toeffect such treatment or prevention for the disease. The“therapeutically effective amount” will vary depending on the compound,the disease and its severity and the age, weight, etc., of the patientto be treated.

When referring to a chemical reaction, the terms “treating”,“contacting” and “reacting” are used interchangeably herein and refer toadding or mixing two or more reagents under appropriate conditions toproduce the indicated and/or desired product. It should be appreciatedthat the reaction which produces the indicated and/or desired productmay not necessarily result directly from the combination of two reagentswhich were initially added, i.e., there may be one or more intermediateswhich are produced in the mixture which ultimately leads to theformation of the indicated and/or desired product.

The term “substantially free of” in reference to a composition, as usedherein, means that the substance form which the composition is free ofcannot be detected by methods known to those skilled in the art.

The term “amorphous” means a material that may be substantially free ofcrystalline impurities, or contain substantial amounts of crystallineimpurities. Amorphous material containing substantial amounts ofcrystalline materials as impurities is hereby referred to as“N-benzoyl-staurosporine essentially amorphous”. As illustrated in FIG.4, the presence of crystalline impurities does not allow for a good haloshape pattern typical of amorphous form that is substantially free ofcrystalline material. The amorphous material that is substantially freeof crystalline forms is hereby referred to as “N-benzoyl-staurosporinepurely amorphous”. FIG. 3 illustrates an X-ray diffractogram pattern forthis form, where the halo shape of the pattern illustrates thesubstantial absence of crystalline structure. Peaks are particularlymissing in the regions characteristics of crystalline form.

The examples further illustrate processes for obtaining both“N-benzoyl-staurosporine essentially amorphous” and“N-benzoyl-staurosporine purely amorphous”.

N-benzoyl-staurosporine is known as(9α,10β,11β,13α)-N-(2,3,10,11,12,13-hexahydro-10-methoxy-9-methyl-1-oxo-9,13,-epoxy-1h,9H-diindolo[1,2,3-gh:3′,2′,1′-1m]pyrrolo[3,4-j][1,7]benzodiazonin-11-yl)-N-methyl-benzamide)and has the following chemical structure:

U.S. Pat. No. 5,093,330 (the '330 patent), incorporated herein byreference in its entirety, claims N-benzoyl-staurosporine. The inventionrelates especially to a particular form preferably that which isreferred to hereinafter as crystalline form II of aN-benzoyl-staurosporine derivative of the compound of formula (I),described above, and to the amorphous forms of N-benzoyl-staurosporine.

Different solid forms of the same drug may exhibit different properties,including characteristics that have functional implications with respectto their use as drug may have substantial differences in suchpharmaceutically important properties as dissolution rates andbioavailability. Likewise, different polymorphs may have differentprocessing properties, such as hydroscopisity, flowability and the like,which could affect their suitability as active pharmaceuticals forcommercial production.

X-ray powder diffraction patterns was measured on a Scintag INC X 1using the Bragg-Brentano parafocusing geometry. The X-ray diffractionpattern depicted in FIG. 1 is summarized in Table 1.

TABLE 1 Powder X-Ray Diffraction Peaks for the Form II CrystalModification of N-benzoyl-staurosporine 2θ (deg) d-spacing (Å) Relativeintensity 3.4 26.1 medium 6.0 14.8 medium 7.8 11.3 medium 8.7 10.1strong 9.2 9.6 medium 9.7 9.1 medium 10.1 8.8 medium 10.4 8.5 low 11.27.9 medium 12.6 7.0 medium 14.1 6.3 medium 15.7 5.7 medium 16.8 5.3medium 18.2 4.9 medium 18.9 4.7 medium 19.3 4.6 medium 19.6 4.5 strong20.2 4.4 strong 24.5 3.6 medium

It should be kept in mind that slight variations in observed 2θ anglesor spacing values are expected based on the specific diffractometeremployed, the analyst and the sample preparation technique. Morevariation is expected for the relative peak intensities. Identificationof the exact crystalline form of a compound should be based primarily onobserved 2θ angles with lesser importance attributed to relative peakintensities.

Some margin of error is present in each of the 2θ angle assignmentsreported herein. The assigned margin of error, in a preferred variant,the crystalline form II of N-benzoyl-staurosporine is approximately ±0.2for each of the peak assignments.

The crystalline form II of N-benzoyl-staurosporine may be alsocharacterized by infrared spectroscopy. The crystalline form II exhibitsa characteristic absorption pattern in infrared (IR) spectroscopicanalysis as depicted in FIG. 2. IR spectroscopic analysis was measuredon a Bruker IFS-55. The crystalline form II of N-benzoyl-staurosporinehas characteristic absorptions, which can be distinguished from that ofother polymorphs, at about 789, 773, 743, 704, 1066, 1026, 1458, 1398,1383, 1602, 1577, 1497, 1627, 1680, 2934 and 3055 cm⁻¹ in IRspectroscopic analysis. Some margin of error is present in each of thecharacteristic absorptions reported herein. The assigned margin of errorin the characteristic absorptions is approximately 2 cm⁻¹ in the rangeof 1900-800 cm⁻¹.

One or more of physical properties and/or spectroscopic properties canbe the basis for characterizing the crystal or polymorphic forms ofN-benzoyl-staurosporine.

The invention also provides a composition containing solidN-benzoyl-staurosporine, which is at least 80%, by total weight of thesolid N-benzoyl-staurosporine in the composition, its crystalline formII. The preferred form of this composition is solidN-benzoyl-staurosporine powder suitable for use as active ingredient informulating pharmaceutical products. The remainder of the solidN-benzoyl-staurosporine in the composition, i.e., 20% or less of thetotal weight of N-benzoyl-staurosporine may be, e.g., other crystallineforms of N-benzoyl-staurosporine. In one specific embodiment, thecomposition contains at least 90% of the crystalline form II ofN-benzoyl-staurosporine with respect to the total weight of the solidN-benzoyl-staurosporine in the composition. In another specificembodiment, the composition contains at least 95% of the crystallineform II with respect to total weight of the solidN-benzoyl-staurosporine in the composition.

In another aspect, the invention also provides processes for purifyingstaurosporine. While the invention is not limited to any specifictheory, the inventors found that a process involves:

-   -   (a) providing a suspension of staurosporine in an alcohol        solvent;    -   (b) contacting the suspension with methanesulfonic acid to get a        solution;    -   (c) further contacting the solution with triethylamine; and    -   (d) isolating the product.

The process further comprises washing the product in a protic or aproticsolvent. A non-limiting example of a protic solvent is ethanol and anon-limiting example of an aprotic solvent is tetrahydrofuran (THF). Theinvention also provides a process for:

-   -   (a) providing a solution of crude staurosporine in either a        protic or aprotic solvent    -   (b) seeding the solution with purified staurosporine;    -   (c) contacting the reaction mixture with an alcohol solvent to        form a precipitate; and    -   (d) isolating the product which is the purified staurosporine.

After the starting material is purified, the material may be convertedto either the crystalline form II or amorphous N-benzoyl-staurosporine.The material may be converted to the crystalline form II ofN-benzoyl-staurosporine, the process comprising:

-   -   (a) reacting staurosporine in an alcohol solvent, such as        ethanol, with benzoic anhydride; and    -   (b) seeding the reaction solution with crystalline form II of        N-benzoyl-staurosporine to produce the product    -   (c) isolating the product which is crystalline form II of        N-benzoyl-staurosporine.    -   The material may be converted to amorphous        N-benzoyl-staurosporine by:    -   (a) reacting staurosporine in a solvent, such as ethanol or THF,        with benzoic anhydride; and    -   (b) seeding the reaction with amorphous N-benzoyl-staurosporine        to produce the product.    -   (c) isolating the product which is amorphous        N-benzoyl-staurosporine.

The product may be further washed with the solvent multiple times toproduce N-benzoyl-staurosporine essentially amorphous.

The technique of spray drying may also be used to prepareN-benzoyl-staurosporine purely amorphous.

The invention also provides a composition containingN-benzoyl-staurosporine essentially amorphous. The invention alsoprovides a composition containing N-benzoyl-staurosporine purelyamorphous.

A process for the preparation of crystalline form II ofN-benzoyl-staurosporine is also provided. The process involves:

-   -   (a) providing a solution of N-benzoyl-staurosporine in either a        protic or aprotic solvent;    -   (b) seeding the solution with crystalline form II of        N-benzoyl-staurosporine;    -   (c) contacting the suspension with an alcohol solvent to form a        precipitate; and    -   (d) isolating the precipitate, which is the crystalline form II        of N-benzoyl-staurosporine.

Non-limiting examples of the protic or aprotic solvents are listed inthe Table below:

Examples Benzyl Alcohol Ethanol Dimethyl Sulfoxide (DMSO) Dimethylformamide (DMF) THF Acetic acid Polyethylene glycol (PEG 200)

The peaks of the X-ray powder diffractogram of crystalline form II thatare of medium to low strength obtained from the above identifiedsolvents can show some variations due to the presence of differentsolvent molecules within the crystal lattice structure.

In one embodiment, N-benzoyl-staurosporine is dissolved in benzylalcohol, precipitated by addition of seeding crystals of the crystallineform II of N-benzoyl-staurosporine and addition of ethanol as theanti-solvent, and crystalline form II of N-benzoyl-staurosporine isobtained. This process is highly-reproducible and the resultingcrystalline product has good filtration. The residual solvent content inB6 is around 4.3% wt/wt of benzyl alcohol and 0.5-1% wt/wt of ethanol.The process is shown schematically as:

The above conditions on the selective preparation of the individualcrystal forms are not conclusive. In general, e.g., it is possible tovary parameters such as the weight ratio of the compound of formula (I)to the solvent and anti-solvent.

Also provided are pharmaceutical compositions containing a crystallineform II or amorphous N-benzoyl-staurosporine and a pharmaceuticallyacceptable carrier. In addition to the active compound, thepharmaceutical composition include one or more pharmaceuticallyacceptable carriers, also known as excipients, which ordinarily lackpharmaceutical activity, but have various useful properties which may,e.g., enhance the stability, sterility, bioavailability and ease offormulation of a pharmaceutical composition. These carriers arepharmaceutically acceptable, meaning that they are not harmful to humansor animals when taken appropriately and are compatible with otheringredients in a given formulation. The carriers may be solid,semi-solid or liquid, and may be formulated with the compound in bulk,but ultimately in the form of a unit-dose formulation, i.e., aphysically discrete until containing a specific amount of activeingredient, such as a tablet or capsule. The pharmaceutical compositionsmay include, in addition to a compound of this invention, one or moreactive pharmaceutical compounds.

The pharmaceutical compositions may be in the form of suspensions,solutions, elixirs, aerosols or solid dosage forms.

The pharmaceutical compositions are contemplated in various formulationssuitable for various modes of administration including, but not limitedto, inhalation, oral, rectal, parenteral (including subcutaneous,intradermal, intramuscular and intravenous), implantable and transdermaladministration. The most suitable route of administration in an givencase depends on the duration of the subject's condition, the length oftreatment desired, the nature and severity of the condition beingtreated, and the particular formulation that is being used. Theformulations may be in bulk or in unit dosage form, and may be preparedby methods well-known in the art for a given formulation.

The amount of active ingredient included in a unit dosage form dependson the type of formulation in which the active ingredient is presented.A pharmaceutical composition will generally contain about 0.1% by weightto about 99% by weight of the active ingredient, preferably about 1% byweight to 50% by weight for oral administration and about 0.2% by weightto about 20% by weight for parenteral administration.

Formulations suitable for oral administration include capsules (hard andsoft), cachets, lozenges, syrups, suppositories and tablets, eachcontaining a predetermined amount of the active compound; as a powder orgranules, as a solution or a suspension in an aqueous or non-aqueousliquid; or as an oil-in-water or water-in-oil emulsion. Suchformulations may be prepared by any suitable method of pharmacy thatincludes the step of bringing into association the active compound and asuitable carrier or carriers. The amount of active ingredient per unitdosage of solid formulations may be as described in prior art forpreparations of N-benzoyl-staurosporine.

In another aspect, the invention also provides methods of treatmentusing the compounds and the pharmaceutical compositions of thisinvention. The compounds and compositions of this invention may beadministered to a subject in an amount effective to be used fortreatment and prevention of the conditions disclosed in the '330 patent.By subject is meant a human or an animal, preferably human. Animalscontemplated by this invention include any animal safely treatable bycompounds of this invention. Most notably, the crystalline form II andamorphous N-benzoyl-staurosporine show high anti-proliferative andanti-tumor activity, as a result of Protein Kinase C (PKC) inhibition,which may be extremely useful for cancer treatment. Moreover, theirhighly selective and potent inhibition of PKC may lead to superiorclinical outcomes for the patient, i.e., delay or suppress diseaseprogression, with equally tolerable regimens. Potential applicationsinclude a variety of solid tumors and more specifically for examplebreast cancer, colon cancer, ovarian cancer and leukemia. In addition,various other indications that may be affected by PKC activity may beeffectively treated by these compositions, including multidrugresistance (MDR), one of the major problems in currently employed cancerchemotherapy and inflammatory diseases in general.

The present invention relates especially to crystalline form II andamorphous N-benzoyl-staurosporine disclosed herein for the treatment ofone of the said diseases or in the preparation of a pharmacologicalagent for the treatment thereof.

The invention relates also to a process for the treatment ofwarm-blooded animals suffering from said diseases, especially a tumordisease, wherein a quantity of the crystalline form II or amorphousN-benzoyl-staurosporine, which is effective against the diseaseconcerned, especially a quantity with anti-proliferative and especiallytumor-inhibiting efficacy, is administered to warm-blooded animals inneed of such treatment. The invention relates moreover to the use of thecrystalline form II or amorphous N-benzoyl-staurosporine for theinhibition of the above-mentioned PKC, or for the preparation ofpharmaceutical compositions for use in treating the human or animalbody, especially for the treatment of a variety of solid tumors and morespecifically, e.g., breast cancer, colon cancer, ovarian cancer andleukemia. Depending on species, age, individual condition, mode ofadministration and the clinical picture in question, effective doses,e.g., daily doses of about 1-2500 mg, preferably 1-1000 mg, especially5-500 mg, are administered to warm-blooded animals of about 70 kg bodyweight.

The invention relates also to pharmaceutical preparations which containan effective amount, especially an effective amount for prevention ortreatment of one of the said diseases, of the crystalline form II oramorphous N-benzoyl-staurosporine of formula (I), together withpharmaceutically acceptable carriers which are suitable for topical;enteral, e.g., oral or rectal; or parenteral administration and may beinorganic or organic and solid or liquid. Especially tablets or gelatincapsules containing the active substance together with diluents, e.g.,lactose, dextrose, sucrose, mannitol, sorbitol, cellulose and/orglycerin; and/or lubricants, e.g., silica, talc, stearic acid or saltsthereof, typically magnesium or calcium stearate; and/or PEG, are usedfor oral administration. Tablets may likewise contain binders, e.g.,magnesium aluminum silicate, starches, typically corn, wheat or ricestarch, gelatin, methylcellulose, sodium carboxymethylcellulose and/orpolyvinylpyrrolidone; and, if so desired, disintegrants, e.g., starches,agar, alginic acid or a salt thereof, typically sodium alginate; and/oreffervescent mixtures, or adsorbents, coloring agents, flavors andsweetening agents. The pharmacologically active compounds of the presentinvention may further be used in the form of preparations for parenteraladministration or infusion solutions. Such solutions are preferablyisotonic aqueous solutions or suspensions, these possibly being preparedbefore use, e.g., in the case of lyophilised preparations containing theactive substance either alone or together with a carrier, e.g.,mannitol. The pharmaceutical substances may be sterilised and/or maycontain excipients, e.g., preservatives, stabilisers, wetting agentsand/or emulsifiers; solubilizers; salts for the regulation of osmoticpressure; and/or buffers. The present pharmaceutical preparations which,if so desired, may contain further pharmacologically active substances,such as antibiotics, are prepared in a manner known per se, e.g., bymeans of conventional mixing, granulating, coating, dissolving orlyophilising processes, and contain from about 1-100%, especially fromabout 1% to about 20%, of the active substance or substances.

The invention is further defined by reference to the following examplesdescribing in detail the preparation of the compound and thecompositions of the present invention, as well as their utility. It willbe apparent to those skilled in the art, that many modifications, bothto materials, and methods, may be practiced with out departing from thepurpose and interest of this invention. The examples that follow are notintended to limit the scope of the invention as defined hereinabove oras claimed below.

EXAMPLES Example 1 Purification of Staurosporine B1 to Staurosporine B2Using Methanesulfonic Acid and Triethylamine

A reactor is charged with staurosporine B1 and ethanol. The suspensionis heated to about 70° C. Methanesulfonic acid is added to thesuspension and the obtained solution is aged. Activated charcoal andHyflo are added and the mixture is aged. The suspension is filtered andthe filter apparatus and filter cake are washed multiple times withethanol. The clear filtered solution is transferred to another reactorand cooled to about 60° C. Triethylamine diluted in ethanol is addedslowly to the reaction mixture. Afterwards, the suspension is cooled toabout 20° C. and aged. The staurosporine B2 is isolated by filtration.The product is rinsed multiple times with ethanol.

Example 2 Purification of Crude Staurosporine B1 to Staurosporine B2Through Crystallization

A reactor is charged with staurosporine B1 and benzyl alcohol. Thesuspension is heated to about 85° C. and aged. Activated charcoal andHyflo are added to the solution and the mixture is aged again. Thesuspension is filtered at about 90° C. The filter apparatus and filtercake are washed multiple times with benzyl alcohol. The clear filteredsolution is transferred to another reactor and cooled to about 70° C.The solution is seeded with staurosporine B2 suspended in ethanol.Afterwards, ethanol is added to the suspension and the suspension isaged, then cooled to about 0° C. The suspension is again aged for atleast 2 hours at about 0° C. The staurosporine B2 is isolated byfiltration. The product is rinsed multiple times with ethanol.

Example 3 Benzylation of Staurosporine B2 to Prepare EssentiallyAmorphous N-benzoyl-staurosporine

A reactor is charged with staurosporine B2 with benzoic anhydride,ethanol and water. The reaction mixture is heated to about 70° C. andaged. The solution is filtered and the filter apparatus and filter cakeare washed multiple times with ethanol. The filtered solution istransferred to another reactor and cooled to about 60° C. Water isslowly added to the solution. The solution is seeded with amorphousN-benzoyl-staurosporine. The suspension is aged, then cooled to about20° C. then aged again. N-benzoyl-staurosporine is isolated byfiltration. The wet cake of N-benzoyl-staurosporine is rinsed multipletimes with absolute ethanol. The wet cake is then re-suspended in thefilter apparatus in ethanol, aged at about 25° C. then cooled to about0-5° C. and aged again. This process step may be repeated multipletimes. N-benzoyl-staurosporine is isolated by filtration. The wet cakeis dried in a vacuum oven at an external temperature of about 40° C. anda vacuum of approximately 10-20 mbar for about 24 hours to obtainproduct: 35-40 g dried cake: 27 g theory quell. Yield: approximately 82%of theory theory quell related to B1 100%.

The use of ethanol in the above reaction may be replaced with THF.

Example 4 Benzylation of Staurosporine B2 to Prepare Crystalline Form IIof N-benzoyl-staurosporine B4

A reactor is charged with staurosporine B2 with benzoic anhydride,absolute ethanol and water. The reaction mixture is heated to about 70°C. and aged. At the end of the reaction time, the reaction mixture isseeded with crystalline form II of N-benzoyl-staurosporine suspended inabsolute ethanol. After aging, absolute ethanol and water are slowlyadded to the suspension. The suspension is aged again, then cooled toabout 0° C. before crystalline form II of N-benzoyl-staurosporine isisolated by filtration. The wet cake of N-benzoyl-staurosporine isrinsed multiple times with absolute ethanol and then dried in a vacuumoven at an external temperature of 60° C. and a vacuum of approximately5-30 mbar for about 16 hours to obtain the product: 39.70 g, dried cake:27.2 g theory quell. Yield: approximately 91.5% of theory quell relatedto B1 100%.

Example 5 Preparation of Crystalline Form II of N-benzoyl-staurosporineUsing Benzyl Alcohol and Ethanol

A reactor is charged with crude N-benzoyl-staurosporine and benzylalcohol. The mixture is heated to 85° C. and the solution issubsequently aged. The solution is cooled to about 70° C. then seededwith purified crystalline form II of N-benzoyl-staurosporine B6suspended in ethanol. Afterwards, ethanol is added and the suspension isaged, then cooled to about 0° C. before crystalline form II ofN-benzoyl-staurosporine is isolated by filtration. The wet cake isrinsed with ethanol multiple times and dried.

Example 6 Preparation of Crystalline Form II of N-benzoyl-staurosporineUsing Acetic Acid and Ethanol

A reactor is charged with crude N-benzoyl-staurosporine and acetic acid.The mixture is heated to about 70° C. and the solution is subsequentlyaged. The solution is then seeded with purified crystalline form II ofN-benzoyl-staurosporine B6 suspended in ethanol. Afterwards, ethanol isadded to the suspension and the suspension is aged, then cooled to about20° C. before crystalline form II of N-benzoyl-staurosporine is isolatedby filtration. The wet cake is rinsed with ethanol multiple times anddried.

Example 7 Preparation of Crystalline Form II Crystal ofN-benzoyl-staurosporine Using PEG

A reactor is s charged with crude N-benzoyl-staurosporine and PEG. Themixture is heated to about 90° C. and the solution is subsequently aged.The solution is cooled to about 70° C. and seeded with purifiedcrystalline form II of N-benzoyl-staurosporine B6 suspended in ethanol.Afterwards ethanol/water in a 1:1 ratio is slowly added and thesuspension is then aged before being cooled to 20° C. The crystallineform II of N-benzoyl-staurosporine is isolated by filtration. The wetcake is rinsed with ethanol multiple times and dried.

Example 8 Preparation of Crystalline Form II of N-benzoyl-staurosporineUsing DMSO

A reactor is charged with N-benzoyl-staurosporine and DMSO. The mixtureis heated to about 70° C. and the solution is subsequently aged thenseeded with purified crystalline form II of N-benzoyl-staurosporine B6suspended in ethanol. Afterwards, ethanol is added to the suspension andthe suspension is aged at 70° C., then cooled to about 20° C. Thesuspension is aged again before crystalline form II ofN-benzoyl-staurosporine is isolated by filtration. The product is rinsedwith ethanol multiple times and dried.

Example 9 Preparation of Crystalline Form II of N-benzoyl-staurosporineUsing DMF

A reactor is charged with N-benzoyl-staurosporine and DMF. The mixtureis heated to about 70° C., aged and then seeded with purifiedcrystalline form II of N-benzoyl-staurosporine B8 in ethanol.Afterwards, ethanol is added to the suspension and the suspension isaged at about 70° C. then cooled to about 20° C. The suspension is againaged before the crystalline form II of N-benzoyl-staurosporine isisolated by filtration. The product is rinsed with ethanol multipletimes and dried.

Example 10 Preparation of Crystalline Form II of N-benzoyl-staurosporineUsing THF

A reactor is charged with amorphous N-benzoyl-staurosporine and THF. Themixture is heated to about 65° C., aged and then seeded with crystallineform II of N-benzoyl-staurosporine B6 in THF. The suspension is aged,then cooled to about 20° C. and again aged before crystalline form II ofN-benzoyl-staurosporine is isolated by filtration. The product is rinsedmultiple times with THF and dried.

Example 11 Preparation of Crystalline Form II of N-benzoyl-staurosporineUsing Ethanol

A reactor is charged with amorphous N-benzoyl-staurosporine and ethanol.The mixture is heated to about 70° C., aged and then seeded withcrystalline form II of N-benzoyl-staurosporine B6 in ethanol. Thesuspension is aged, the cooled to about 20° C. and again aged beforecrystalline form II of N-benzoyl-staurosporine is isolated byfiltration. The product is rinsed multiple times with ethanol and dried.

Example 12 Spray Drying Experiment to Get Amorphous PKC412

The feed solution is prepared by dissolving 10 g of PKC412 (crystallineform II) in approximately 160 g of dichloromethane. The feed is ready tobe atomized into droplets from which the organic solvent is evaporatedin the drying chamber by heated nitrogen resulting in the formation ofamorphous PKC412. The inlet temperature is set to 60° C. whereas theoutlet temperature is set to 40° C. The residual solvent content afterspray drying was approximately 1% w/w. PKC412 is further dried undervacuum to get a solvent free material.

Example 13 Of Spray Drying Experiment to Get Amorphous PKC412

The feed solution was prepared by dissolving 7 g of PKC412 (crystallineform II) in approximately 400 g of THF. The feed is ready to be atomizedinto droplets from which the organic solvent is evaporated in the dryingchamber by heated nitrogen resulting in the formation of amorphousPKC412. The inlet temperature is set to 110-120° C. whereas the outlettemperature is set to 70-80° C. The residual solvent content after spraydrying is approximately 3% w/w. PKC412 is further dried under vacuum toget a solvent free material.

Example 14 Of Spray Drying Experiment to Get Amorphous PKC412

The feed solution is prepared by dissolving 7 g of PKC412 (crystallineform II) in approximately 350 g of ethanol/acetic acid 70:30 w/w. Thefeed is ready to be atomized into droplets from which the organicsolvent is evaporated in the drying chamber by heated nitrogen resultingin the formation of amorphous PKC412. The inlet temperature is set to215° C. whereas the outlet temperature is set to 120° C. The residualsolvent content after spray drying is below 0.5%. Further conventionaldrying is not required.

Example 15 Tablets with Crystalline Form II of N-benzoyl-staurosporine

Tablets containing 100 mg of the active substance named in the title areusually prepared in the following composition:

Composition Amount (mg) Active ingredient 100 Crystalline lactose 240Avicel 80 PVPPXL 20 Aerosil 2 Magnesium stearate 5 Total 447

Preparation: The active substance is mixed with carrier materials andcompressed on a tableting machine (Korsch EKO, punch diameter 10 mm).

Avicel is microcrystalline cellulose (FMC, Philadelphia, USA).

PVPPXL is polyvinylpolypyrrolidone, cross-linked (BASF, Germany).

Aerosil is silicon dioxide (Degussa, Germany).

Example 16 Capsules with Crystalline Form II of N-benzoyl-staurosporine

Capsules containing 100 mg of the compound named in the title as activesubstance are usually prepared in the following composition:

Composition Amount (mg) Active ingredient 100 Avicel 200 PVPPXL 15Aerosil 2 Magnesium stearate 1.5 Total 318.5

The capsules are prepared by mixing the components and filling themixture into hard gelatin capsules, size 1.

1. A process for purifying staurosporine comprising: (a) providing asuspension of staurosporine in an alcohol solvent; (b) contacting thesuspension with methanesulfonic acid; (c) further contacting thesolution with triethylamine; and (d) isolating the product.
 2. Theprocess according to claim 1, further comprising washing the isolatedproduct with the alcohol solvent.
 3. The process according to claim 1,wherein the alcohol solvent is ethanol.
 4. A process for purifyingstaurosporine comprising: (a) providing a solution of staurosporine in afirst solvent; (b) seeding the solution with purified staurosporine in asecond solvent; and (c) isolating the product.
 5. The process accordingto claim 4, wherein the first solvent is selected from benzyl alcohol,DMF, DMSO, acetic acid or PEG.
 6. The process according to claim 4,wherein the second solvent is selected from ethanol, THF or a solutionof ethanol in water.
 7. A process for the preparation ofN-benzoyl-staurosporine comprising: (a) reacting staurosporine withbenzoic anhydride to form a solution; (b) seeding the solution withamorphous N-benzoyl-staurosporine; and (c) isolating the product.
 8. Acompound which is amorphous N-benzoyl-staurosporine produced by theprocess of claim
 7. 9. The process according to claim 7, furthercomprising spray drying the product.
 10. A compound which is amorphousN-benzoyl-staurosporine.
 11. The compound according to claim 10, havingsubstantially the same X-ray diffraction pattern as shown in FIG.
 3. 12.The compound according to claim 10, having substantially the same X-raydiffraction patter as shown in FIG.
 4. 13. A pharmaceutical compositioncomprising: (a) the compound of claim 10; and (b) a pharmaceuticallyacceptable carrier or diluent.
 14. The pharmaceutical compositionaccording to claim 13, further comprising one or more pharmaceuticallyacceptable excipients.
 15. The pharmaceutical composition according toclaim 13, is in a dosage form suitable for oral administration.
 16. Thepharmaceutical composition according to claim 10, wherein the dosageform is selected from a tablet, capsule or solution.
 17. Use ofamorphous N-benzoyl-staurosporine according to claim 10, for thepreparation of a pharmacological agent for the treatment of a tumourdisease.